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Literature summary for 3.6.5.3 extracted from

  • De Laurentiis, E.I.; Wieden, H.J.
    Identification of two structural elements important for ribosome-dependent GTPase activity of elongation factor 4 (EF4/LepA) (2015), Sci. Rep., 5, 8573.
    View publication on PubMedView publication on EuropePMC

Activating Compound

Activating Compound Comment Organism Structure
50S ribosome stimulates the enzyme Escherichia coli
70S ribosome required, stimulates the enzyme Escherichia coli
additional information the 30S ribosome does not stimulate the enzyme Escherichia coli

Cloned(Commentary)

Cloned (Comment) Organism
gene lepA, recombinant overexpression of His-tagged wild-type and mutant enzymes in Escherichia coli strain BL21(DE3) Escherichia coli

Protein Variants

Protein Variants Comment Organism
H81A site-directed mutagenesis Escherichia coli
additional information generation of C-terminal domain truncation mutants of the enzyme, truncation of the C-terminal domain does not abolish ribosome binding, the mutant enzyme is stabilized on the ribosome when the C-terminal domain is removed. Binding of wild-type EF4 and mutant variants to the ribosome in the presence of guanine nucleotides, kinetics and affinities Escherichia coli

Inhibitors

Inhibitors Comment Organism Structure
Thiostrepton inhibits the stimulation of the enzyme's GTPase activity by the 70S ribosome Escherichia coli

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
additional information
-
additional information Michaelis-Menten kinetics of GTP hydrolysis, stopped flow measurements, kinetic analysis of wild-type and mutant enzymes, overview Escherichia coli
0.00032
-
GTP recombinant wild-type enzyme in presence of 70S ribosome, pH 7.5, 37°C Escherichia coli
0.27
-
GTP recombinant wild-type enzyme in absence of 70S ribosome, pH 7.5, 37°C Escherichia coli

Metals/Ions

Metals/Ions Comment Organism Structure
Mg2+ required Escherichia coli

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
GTP + H2O Escherichia coli
-
GDP + phosphate
-
?

Organism

Organism UniProt Comment Textmining
Escherichia coli P60785 gene lepA
-

Purification (Commentary)

Purification (Comment) Organism
recombinant His-tagged wild-type and mutant enzymes from Escherichia coli strain BL21(DE3) by nickel affinity chhrmatography and gel filtration Escherichia coli

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
GTP + H2O
-
Escherichia coli GDP + phosphate
-
?
additional information enzyme-ribosome binding analysis, overview. Binding of wild-type EF4 and mutant variants to the ribosome in the presence of guanine nucleotides, kinetics and affinities Escherichia coli ?
-
?

Synonyms

Synonyms Comment Organism
EF4
-
Escherichia coli
elongation factor 4
-
Escherichia coli
LepA
-
Escherichia coli
ribosome-dependent GTPase
-
Escherichia coli

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
37
-
assay at Escherichia coli

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
0.003
-
GTP recombinant wild-type enzyme in absence of 70S ribosome, pH 7.5, 37°C Escherichia coli
2.3
-
GTP recombinant wild-type enzyme in presence of 70S ribosome, pH 7.5, 37°C Escherichia coli

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
7.5
-
assay at Escherichia coli

General Information

General Information Comment Organism
malfunction successive removal of the C-terminus impairs ribosome-dependent multiple turnover GTPase activity of enzyme EF4 Escherichia coli
additional information the last 44 C-terminal amino acids of elongation factor 4 form a subdomain within the C-terminal domain that is important for GTP-dependent function on the ribosome. Efficient nucleotide hydrolysis by the enzyme on the ribosome depends on its conserved residue His 81, which is essential for catalysis in EF4 Escherichia coli

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
0.007
-
GTP recombinant wild-type enzyme in presence of 70S ribosome, pH 7.5, 37°C Escherichia coli
0.011
-
GTP recombinant wild-type enzyme in absence of 70S ribosome, pH 7.5, 37°C Escherichia coli